Molecular Epidemiology of Aspergillus fumigatus Isolates Harboring the TR ₃₄ /L98H Azole Resistance Mechanism

Abstract

A rapid emergence of azole resistance has been observed in Aspergillus fumigatus in The Netherlands over the past decade. The dominant resistance mechanism appears to be of environmental origin and involves the TR 34 /L98H mutations in cyp51A. This resistance mechanism is now also increasingly being found in other countries. Therefore, genetic markers were used to gain more insights into the origin and spread of this genotype. Studies of 142 European isolates revealed that those with the TR 34 / L98H resistance mechanism showed less genetic variation than azole-susceptible isolates or those with a different genetic basis of resistance and were assigned to only four CSP (putative cell surface protein) types. Sexual crossing experiments demonstrated that TR 34 /L98H isolates could outcross with azole-susceptible isolates of different genetic backgrounds, suggesting that TR 34 / L98H isolates can undergo the sexual cycle in nature. Overall, our findings suggest a common ancestor of the TR 34 /L98H mechanism and subsequent migration of isolates harboring TR 34 /L98H across Europe.A spergillus fumigatus is a saprophytic fungus that is capable of causing a wide range of diseases in various hosts. Invasive aspergillosis is the most severe manifestation of Aspergillus infection in humans, and this disease is associated with substantial mortality and morbidity. Medical triazoles, such as itraconazole, voriconazole, and posaconazole, play an important role in the management of Aspergillus diseases. However, azole resistance is an emerging problem in A. fumigatus and has been shown to be associated with increased probability of treatment failure (10,11,19,20,30,35,37,39,41).Azole resistance is commonly due to mutations in the cyp51A gene, which encodes 14-␣-demethylase in the ergosterol biosynthesis pathway. In azole-resistant clinical A. fumigatus isolates, a wide variety of cyp51A mutations, such as substitutions at codons G54, G138, P216, F219, M220, and G448, have been found (5,11,29). This is in contrast with a different pattern of resistance observed in isolates from The Netherlands. Here, a resistance mechanism consisting of the L98H substitution together with a 34-bp tandem repeat (TR 34 ) in the promoter region of this gene (TR 34 / L98H) was found to be present in over 90% of itraconazole-resistant isolates, which also showed reduced susceptibility to voriconazole and posaconazole (30,36). TR 34 /L98H isolates were recovered primarily from azole-naïve patients and were also recovered from the environment (28, 36). These observations suggest that azole-resistant Aspergillus is acquired by patients from an environmental source rather than arising through azole therapy. Recently, we provided evidence that exposure of A. fumigatus to 14-␣-demethylase inhibitor (DMI) fungicides might provide a selective pressure leading to the emergence of TR 34 /L98H resistant isolates in the environment (27). On the basis of in vitro crossresistance, molecule alignment studies, and docking simulations, five triazole fungicides that ...

Multiple cyp51A -Based Mechanisms Identified in Azole-Resistant Isolates of Aspergillus fumigatus from China

“…Four isolates were identified as azole resistant from a total of 72 A. fumigatus isolates, of which three are highly resistant to ITR and harbor the TR34/L98H mutation that was described previously by Lockhart et al (29) In their ARTEMIS study, a total of eight azole-resistant isolates (itraconazole MIC of Ն16 mg/liter) were recovered from Hangzhou, and all exhibited the TR34/L98H mutation together with S297T and F495I. As did Lockhart et al, we found that the TR34/L98H mutation is still a predominant mutation which is quite common in Europe and some other Asian countries (10,15,17,18,20,36,37). Some evidence suggests that this resistant genotype is likely to have arisen as a side effect of widespread use of agricultural fungicides.…”

Epidemiology and Molecular Characterizations of Azole Resistance in Clinical and Environmental Aspergillus fumigatus Isolates from China

“…Increasing evidence indicated that A. fumigatus should be able to generate resistance to antifungal drugs via the sexual cycle to allow evolution in response to environmental change (40,43). However, a linkage disequilibrium analysis study from the Netherlands indicated that all the azole-resistant A. fumigatus isolates nested within a single, predominantly asexual, population (44).…”

In Vitro Biochemical Study of CYP51-Mediated Azole Resistance in Aspergillus fumigatus

“…The five A. fumigatus CYP51A positions, or hot spots, most frequently undergoing mutations responsible for conferring azole resistance are glycine-54, leucine-98, glycine-138, methionine-220, and glycine-448 (4). G54, G138, M220, and G448 CYP51A point mutations are thought to have arisen during triazole therapy of patients in the clinic (5,6), while TR 34 /L98H and TR 46 /Y121F/T289A may have arisen in the environment in the Netherlands in response to the use of agricultural triazole fungicides (7)(8)(9)(10).…”